Purification, identification, and characterization of two GTP-binding proteins with molecular weights of 25,000 and 21,000 in human platelet cytosol. One is the rap1/smg21/Krev-1 protein and the other is a novel GTP-binding protein.

摘要: Abstract We have purified, characterized, and identified two GTP-binding proteins with Mr of 25,000 (c25KG) 21,000 (c21KG) from the cytosol fraction human platelets. These were not copurified beta gamma subunits heterotrimeric proteins. Amino acid sequences tryptic fragments c21KG completely matched those rap1 protein (Pizon, V., Chardin, P., Lerosey, I., Olofsson, B., Tavitian, A. (1988) Oncogene 3, 201-204), smg p21 (Kawata, M., Matsui, Y., Kondo, J., Hishida, T., Teranishi, Takai, Y. J. Biol. Chem. 263, 18965-18971), Krev-1 (Kitayama, H., Sugimoto, Matsuzaki, Ikawa, Noda, M. (1989) Cell 56, 77-84). The partial amino sequence analysis c25KG revealed that this was different any low already reported. bound about 1 mol [35S] guanosine 5'-(3-O-thio)triphosphate (GTP S)/mol protein, a Kd value 45 nM. [35S]GTP S-binding to specifically inhibited by guanine nucleotides, GTP GDP, but adenine nucleotides such as ATP adenyl-5'-yl beta, gamma-imidodiphosphate. binding activity pretreatment N-ethylmaleimide. hydrolyzed librate Pi specific 1.8 mmol Pi/mol protein/min, which are activities purified conclude is novel rap1/smg p21/Krev-1 product.